An activatable fluorescence probe for rapid detection and in situ imaging of ß-galactosidase activity in cabbage roots under heavy metal stress.

ß-Galactosidase (ß-gal), an enzyme related to cell wall degradation, plays an important role in regulating cell wall metabolism and reconstruction. However, activatable fluorescence probes for the detection and imaging of ß-gal fluctuations in plants have been less exploited. Herein, we report an activatable fluorescent probe based on intramolecular charge transfer (ICT), benzothiazole coumarin-bearing ß-galactoside (BC-ßgal), to achieve distinct in situ imaging of ß-gal in plant cells. It exhibits high sensitivity and selectivity to ß-gal with a fast response (8 min). BC-ßgal can be used to efficiently detect the alternations of intracellular ß-gal levels in cabbage root cells with considerable imaging integrity and imaging contrast. Significantly, BC-ßgal can assess ß-gal activity in cabbage roots under heavy metal stress (Cd2+, Cu2+, and Pb2+), revealing that ß-gal activity is negatively correlated with the severity of heavy metal stress. Our work thus facilitates the study of ß-gal biological mechanisms.

Fluorescent Probe for Investigating the Mitochondrial Viscosity and Hydrogen Peroxide Changes in Cerebral Ischemia/Reperfusion Injury.

Cerebral ischemia-reperfusion injury (CIRI), a cause of cerebral dysfunction during cerebral infarction treatment, is closely associated with mitochondrial viscosity and hydrogen peroxide (H2O2). However, the accurate measurement of mitochondrial viscosity and H2O2 levels in CIRI is challenging because of the lack of sufficient selectivity and blood-brain barrier (BBB) penetration of existing monitoring tools related to CIRI, hampering the exploration of the role of mitochondrial viscosity and H2O2 in CIRI. To address this issue, we designed an activatable fluorescent probe, mitochondria-targeting styryl-quinolin-ium (Mito-IQS), with excellent properties including high selectivity, mitochondrial targeting, and BBB penetration, for the visualization of mitochondrial viscosity and H2O2 in the brain. Based on the real-time monitoring capabilities of the probe, bursts of mitochondrial viscosity and H2O2 levels were visualized during CIRI. This probe can be used to monitor the therapeutic effects of butylphthalein treatment. More importantly, in vivo experiments further confirmed that CIRI was closely associated with the mitochondrial viscosity and H2O2 levels. This discovery provides new insights and tools for the study of CIRI and is expected to accelerate the process of CIRI diagnosis, treatment, and drug design.

A large Stokes shift NIR fluorescent probe for visual monitoring of mitochondrial peroxynitrite during inflammation and ferroptosis and in an Alzheimer's disease model.

The excessive formation of peroxynitrite (ONOO-) in mitochondria has been implicated in various pathophysiological processes and diseases. However, owing to short emission wavelengths and small Stokes shifts, previously reported fluorescent probes pose significant challenges for mitochondrial ONOO- imaging in biological systems. In this study, a near-infrared (NIR) fluorescent probe, denoted as DCO-POT, is designed for the visual monitoring of mitochondrial ONOO-, displaying a remarkable Stokes shift of 170 nm. The NIR fluorophore of DCO-CHO is released by DCO-POT upon the addition of ONOO-, resulting in off-on NIR fluorescence at 670 nm. This phenomenon facilitates the high-resolution confocal laser scanning imaging of ONOO- generated in biological systems. The practical applications of DCO-POT as an efficient fluorescence imaging tool are verified in this study. DCO-POT enables the fluorometric visualization of ONOO- in organelles, cells, and organisms. In particular, ONOO- generation is analyzed during cellular and organism-level (zebrafish) inflammation during ferroptosis and in an Alzheimer's disease mouse model. The excellent visual monitoring performance of DCO-POT in vivo makes it a promising tool for exploring the pathophysiological effects of ONOO-.

A turn-on NIR fluorescent probe for risk-assessing oxidative stress in cabbage roots under abiotic stress.

Oxidative stress is closely related to the crop health status under stress conditions. H2O2 is an important signaling molecule in plants under stress. Therefore, monitoring H2O2 fluctuations is of great significance when risk-assessing oxidative stress. However, few fluorescent probes have been reported for the in situ tracking of H2O2 fluctuations in crops. Herein, we designed a "turn-on" NIR fluorescent probe (DRP-B) to detect and in situ-image H2O2 in living cells and crops. DRP-B exhibited good detection performance for H2O2 and could image endogenous H2O2 in living cells. More importantly, it could semi-quantitatively visualize H2O2 in cabbage roots under abiotic stress. Visualization of H2O2 in cabbage roots revealed H2O2 upregulation in response to adverse environments (metals, flood, and drought). This study provides a new method for risk-assessing oxidative stress in plants under abiotic stress and is expected to provide guidance for the development of new antioxidant defense strategies to enhance plant resistance and crop productivity.